ABSTRACT
OBJECTIVE To investigated the regulatory effect of paeoniflorin-6'-O-benzene sulfonate (CP-25) on B cell activating factor (BAFF)/BAFF receptor-nuclear factor of kappa B (NF-κB) signaling in B cell of collagen induced-arthritis (CIA) mice. METHODS Mice CIA was induced by injection of typeⅡcollagen (CⅡ). The arthritis index (AI) and swollen joint count (SJC) were assessed, and histopathology of spleen and joints were observed. The percentage of B cells subsets, BAFF receptor expressions were analyzed by flow cytometry. BAFF and immunoglobulin (Ig) levels were measured by protein antibody array. The expressions of TRAF2, MKK3, MKK6, p-P38, and p-NF-κB65 in NF-κB signaling mediated by BAFF were analyzed by western blot. RESULTS CP-25 decreased AI and SJC, restored abnormal weights, reduced thymus index and spleen index, inhibited T/B cells proliferation, alleviated the histopathology of spleen and joints in CIA mice. CP-25 also reduced high levels of serum BAFF and immunoglobulin, decreased CD19+B cells, CD19+CD27+B cells, and CD19-CD27+CD138+ plasma cells, inhibited BAFFR and TACI expressions, decreased the expressions of TRAF2, MKK3, MKK6, p-P38, and p-NF-κB65. Compared with biological agents etanercept and rituximab, CP-25 restored high T cells proliferation and percentages of B subsets to normal level, and recovered the high levels of IgA, IgD, IgG1, IgG2a and high expressions molecules in NF- κB signaling to normal levels. The action intensity of rituximab and etanercept was more strong than CP- 25. The inhibitor effects of rituximab and etanercept on AI and SJC, thymus index, proliferation of T cells and B cells subsets were strong, and down-regulated the indexes to under normal levels. CONCLUSION CP-25 might be a promising anti- inflammatory immune and regulation drug, which alleviated CIA and regulated the functions of B cells through BAFF/BAFF receptor-NF-κB signaling.
ABSTRACT
OBJECTIVE This study was to investigate the effects of CP- 25 on the functions of activated human B cells through regulating BAFF and TNF-alpha signaling. METHODS B cells from peripheral blood mononuclear cells (PBMCs) of normal human were isolated using magnetic cell separation (MACS) by a positive selection. B cells (107 cells·mL-1) were stimulated by BAFF (100 ng·mL-1) or TNF-alpha (100 ng·mL-1) for two hours, and then were treated with CP-25 (10-5 mol·L-1) or Rituximab (5 μg·mL-1) or Etanercept (10 μg·mL-1). B cell proliferation was detected by CCK-8. B cell subsets and BAFF receptors (BAFFR, BCMA and TACI) were analyzed by flow cytometry. The expression of TNFR1 and TNFR2 on B cells was analyzed by flow cytometry. The expression of MKK3, MKK6, P-p38, P-p65, TRAF2 and p100/52 was analyzed by Western blotting. RESULTS CP-25 inhibited B cells proliferation stimulated by BAFF or TNF- alpha. CP- 25, Rituximab and Etanercept reduced the percentage and numbers of CD19+ B cells, CD19+CD20+ B cells, CD19+CD27+ B cells and CD19+CD20+CD27+ B cells induced by BAFF or TNF-alpha. CP-25 down-regulated the high expression of BAFFR, BCMA and TACI stimulated by BAFF or TNF-alpha. CP-25, Rituximab and Etanercept down-regulated significantly the expression of TNFR1 and TNFR2 on B cell stimulated by BAFF or TNF-alpha. CP-25, Rituximab and Etanercept down-regulated the expression of MKK3, P-p38, P-p65, TRAF2 and p52 in B cells stimulated by BAFF and the expression of TRAF2 and P- p65 in B cells stimulated by TNF-alpha. CONCLUSION CP- 25 regulated moderately activated B cells function by by regulating the classical and alternative NF-κB signaling pathway mediated by BAFF and TNF-alpha-TRAF2-NF-κB signaling pathway. This study suggests that CP-25 may be a promising anti-inflammatory immune and soft regulation drug.